David Baram from Clifton Enterprises Sampling in the Bioprocessing Industries: Expectations and the...

David Baram from Clifton Enterprises

Sampling in theSampling in the Bioprocessing Industries: Bioprocessing Industries:

Expectations andExpectations andthe True Picturethe True Picture

Presented by:David Baram from Clifton Enterprises

Sheffield, Englandemail: [email protected]

Interphex Japan

בס"ד


IntroductionIntroduction• In the Bioprocessing Industry, sampling is fundamental. However,

since people have been doing it for years, it has become a routine. Unfortunately, when things become routine, people start to overlook the related pitfalls and the possible detrimental effect on their processes as a whole.

• This presentation, presented by Clifton Enterprises, is designed to review the various stages of sampling and the potential negative consequences associated with each one of those stages. This presentation will introduce state of the art technology, developed with the support of the British Government through the Department of Trade and Industry, to eliminate these consequences.

• Before I continue, I would like to take this moment to thank Sei Murakami from Hitachi Industrial System Div. for giving me this opportunity to be here today.


The Purpose of SamplingThe Purpose of Sampling

• Monitor the Process For example, by checking:

» pH

» Growth Rate

» Glucose

• Identify Contamination

• Validation

Importance in Choosing Right“The sampling system is responsible for collecting a representative sample of the process stream and delivering it to the analyzer for analysis.

Obviously, the reliability of the sampling system directly affects the reliability of the overall composition analysis system.

The transport delay associated with the sampling system contributes directly to the overall deadtime for an on-line composition measurement. For example, an improperly designed sampling system can result in a transport time of one hour … Sampling deadtime can have a dramatic effect on the performance …”

Chemical and Bio-Process Control (Third edition)

By James B. Riggs and M. Nazmul Karim



Components in Sterile SamplingComponents in Sterile Sampling• Sterile access to the process fluid

– Normally via a Sampling Valve

• Collection System– Open– Bottle– Bag

• Access to Sample for Analysis– Normally via a Syringe


Issues with Common Sampling Issues with Common Sampling ValvesValves

• Mechanism

• Steam Sterilization

• CIP

• Internal Volume

• Dead Volume

• Maintenance


Common Sampling Valve

Problem Areas

Plug & Seat Elastomeric Material Temp. & Chemical

Limitations

Cleanability

& Sterility

Volume Operating Mechanism

Design of Common Sampling Valves


Issues with Common Collecting Issues with Common Collecting EquipmentEquipment

• Complex operation– Difficult to maintain sterility

• Intricate human interaction process– Prone to Human Error

• Requires Wasteful Sterilization Flush

• Lengthy procedure– Detrimental for Ensuring Viable Sample


A typical 4-valve rig for two bottles sampling

• Autoclave a bagged sample bottle with a filter and a valve (V3) sealing the opening.• Unbag the sample bottle as shown, connecting the closed V3 to the closed Tank Sample Valve V2.• Steam behind the flow paths of V2 and V3 as shown in pink.• Close Steam source (V1) and Condensate Drain (V4). You now have a sterile path between your

process and your sample bottle.• Open the Sample bottle (V3) and the Tank Sample Valve (V2), and take your sample as shown in blue.• Close the Sample Valve (V2), then the Sample receptacle Valve (V3). Drain V4 and remove your

Sample Bottle.


Issues with Accessing Issues with Accessing Samples in Bottles and Common BagsSamples in Bottles and Common Bags

• Requires Safety Cabinet

• Requires replacing a Bottle with a Septum– Risks contamination– Time Consuming

• Sample withdrawal requires syringe with needle– Needle-stick risk to personnel– Insurance factor

• Time required compromises sample viability– pH change– Contamination


• No Maintenance

• Cleanability assured (No Cross Contamination)

• Continuous Steam Sterilization, if required

• No Dead Volume

• Minimal Internal Volume

• 3-Position Valve


Simple 3-Position Mechanism

Sterilize

Shutoff

Sample


Vanasyl Sampling Valve with Vanasyl Sampling Valve with Bottles SamplingBottles Sampling


Vanasyl Sampling Valve with Bottles Sampling


Areas for Bag Technology Applications

• Bioreactor sampling

• Bioreactor additions (antifoam etc.)

• Media/buffer prep & storage

• Product storage & transfer

• Lab & pilot scale bioreactors

• Aseptic connections


Examples of Common Sampling Bags


Example Sampling Bags Assembly


Technical Data for Ethylene-Vinyl Acetate (EVA)(Film Gauge 350 μm @ 232)

Physical Properties

Dimension Value Procedure

Water Vapour Transmission

Rate

g/(m2.day) 4 ASTM F-1249

Oxygen Permeability

cm2/(m2.day.bar) 800 ASTM D-3985

Carbon Dioxide Permeability

cm2/(m2.day.bar) 7000 Mocon Permatran C-IV


Technical Data for Multilayer Ethylene-vinylacetate (EVA) with Barrier(Film Gauge 400 μm @ 232)

EVA Without a Barrier

Physical Properties

Dimension Value Procedure

Water Vapour Transmission

Rate

g/(m2.day) 1.644

ASTM F-1249

Oxygen Permeability cm2/(m2.day.bar) 2.2800

ASTM D-3985

Carbon Dioxide Permeability

cm2/(m2.day.bar) 67000

Mocon Permatran C-IV

UV Transmission


Comparison Between Bottles and the Vanasyl Sampling BagBag vs. Bottle Samples

0

5

10

15

20

25

0 5 10 15 20 25 30 35 40 45 50

Run time

Glu

cose o

r pC

O2 (%

)

BagPCO2

BagGluc.

BottlePC02

BottleGluc.

Bag vs. Bottle pH

7.06

7.08

7.1

7.12

7.14

7.16

7.18

7.2

7.22

7.24

0 5 10 15 20 25 30 35 40 45 50

Time (hrs.)

pH BagpH

BottlepH


The Vanasyl Sampling BagThe Vanasyl Sampling Bag


Sampling Process Flow - Bottle vs. BagCOMMON SAMPLING BAGS SYSTEM VANASYL SAMPLING BAGS SYSTEM

1

2 Operator connects the Sampling Bags to the Operator connects the Sampling Bag to thespecific fermentor specific fermentor and draws a sample

3 Eliminated Eliminated

4 Eliminated

5 Eliminated

6 Eliminated

7

8

9 Eliminated

10 Eliminated

Operator collects 2 Sampling Bag which is kept close by. Or a few are connected to the vessel.

Process material (in the majority of Bag Sampling systems) is run into first bag to purge the bags' tube connections &

Second (or first) bag is filled with sample material

Bag with the sample is placed in a Safety Cabinet where the sample is transferred to a bottle which is then sealed with septum cap.

Process material is run into the first bottle to purge the tree.

Bottles are sterilized, capped, and stored or redistributed to various departments.

Bottles are sterilized, capped, and stored or redistributed to various departments.

Bottle is disassembled, prepped and cleaned.Bottles are disassembled, prepped and cleaned.

Bottle with the sample is placed in a Safety Cabinet where the bottle valve cap is replaced with a septum cap.

Operator collects 1 Sampling Bag which is kept close by

Second bottle is filled with sample material

Operator connects the Sampling Bottles to a specialized sample "tree" attached to the Sample Valve for a specific fermentor.

CURRENT SAMPLE BOTTLE PROCESS

Bottles are steamed for a half hour and allowed to cool with one bottle open to avoid forming a vacuum and to allow condensate to leave sample

Operator collects 2 Sterile Sampling Bottles from the Supply which is kept in cabinets, ready for use

The bag with the sample can either be kept frozen for validation or discarded.

The Sampling Bag is labelled and sent for analysis where the samples are drawn via a syringe (without a needle), which is directly connected to the Vanasyl Sampling Bag.

Bottles are removed and the second bottle is labelled and sent for analysis where the samples are drawn through the septum with a needle attached to a syringe.

Bottle with the sample is labelled and sent for analysis where the samples are drawn through the septum with a needle attached to a syringe.

The bags are discarded. The bottle with sample material that if not required to be kept frozen for validation, is rinsed and placed in a bucket for pickup by Glassware's

First bottle with purge material, and second bottle with sample material that if not required to be kept frozen for validation, are rinsed and placed in a bucket for pickup by Glassware's


A Year’s Cost Comparison Between Bottle Sampling and

Vanasyl Bag Sampling (True in 2013)

Number of Samples/year on 14 Fermenters 11,814

Process Bottles/year 23,628

Sampling Bags/year 11,814

Estimated Bottle Sampling Costs

Req. Material for Maintaining the Valves and Bottle (Replacement Bottles, Filters, Diaphragms, Replacement Valves, etc.) $308,960.00

Maintenance Labour Costs (1,656 hours for the bottles and 504 hours on valves @ $40/hour (technician)) $86,400.00

Labour Costs Associated with Sample Taking (11,814 samples, 30 Minutes per Sample @ $70/hour - scientist) $413,490.00

Bottle Sampling Estimated Total Costs $808,850.00

Single Bottle Sampling Estimated Costs $68.47

Estimated Vanasyl Bag Sampling Costs

Bags Costs (11,814 Sample Bags using 150 ml @ $36 each) $425,304.00

Req. Material for Maintaining the Valves and Bottle (O-Rings for the Ingold, Inlet, and Outlet fittings) $300.00

Maintenance Labour Costs (70 Hours @ $40/hour - Technician) $2,800.00

Labour Costs Associated with Sample Taking (11,814 samples, 7 Minutes per Sample @ $70/hour - scientist) $96,481.00

Bags Sampling Estimated Total Costs $524,885.00Single Bag Sampling Estimated Costs $44.43

ESTIMATED YEARLY SAVINGS - GLASS BOTTLES Vs VANASYL BAGS $283,965.00

David Baram


Further Advantages of the Vanasyl Sampling System

• The Vanasyl Sampling System offers 4 easy sampling steps instead of 10 complicated sampling steps as well as:

– More than 99% reduction in product loss. (See Table above)

– Eliminates introduction of steam into process and sample.

– Eliminates false samples due to cross contamination.

– Eliminate the dilution of sample with condensate

• No delays between collection and testing ensuring sample integrity.

• No need for a safety cabinet.

• No use of needles or sharps (eliminate needle injuries and disposal issues).

• Sample can be frozen in the bag to -120C.

Bottle Sampling: A Minimum of 250 ml of product flashing per sample.

A yearly product loss from 11,814 samples: 2,953.50 Litres/year

Vanasyl Bag Sampling: A Maximum 2 ml of product flashing per sample.

A yearly product loss from 11,814 samples: 23.60 Litres/year


Vanasyl Bag Sampling


Vanasyl Sampling Bag System


Clave Connection on Vanasyl Sampling Bag


ConclusionConclusion

• This presentation should raised the awareness of sampling and its related issues and hopefully was beneficial for all those who attended.

David Baram from Clifton Enterprises Sampling in the Bioprocessing Industries: Expectations and the...

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